Paul W. Chrystal, Nils J. Lambacher, Lance P. Doucette, James Bellingham, Elena R. Schiff, Nicole C.L. Noel, Chunmei Li, Sofia Tsiropoulou, Geoffrey A. Casey, Yi Zhai, Nathan J. Nadolski, Mohammed H. Majumder, Julia Tagoe, Fabiana D’Esposito, Maria Francesca Cordeiro, Susan Downes, Jill Clayton-Smith, Jamie Ellingford, Genomics England Research Consortium, Omar A. Mahroo, Jennifer C. Hocking, Michael E. Cheetham, Andrew Webster, Gert Jansen, Oliver E. Blacque, W. Ted Allison, Ping Yee Billie Au, Ian M. MacDonald, Gavin Arno, Michel R. Leroux. “The inner junction protein CFAP20 functions in motile and non-motile cilia and is critical for vision“. Nat Commun (2022), 13,
Identifying a gene spanning cilia function and critical for vision
Cilia are organelles present in most eukaryotic cells and are categorized into discrete groups: motile cilia (such as those found on spermatozoa) are required for locomotion and fluid dynamics whereas non-motile cilia (like those found in photoreceptors of the eye) are required for cell signalling. Paul William Chrystal, post-doctoral fellow at the University of Alberta, led a study that demonstrated that CFAP20 breaks this dichotomy by functioning in BOTH motile and non-motile cilia. They also identified CFAP20 as a novel cause of childhood blindness in humans, increasing the list of therapeutic targets for patients with the disease.
CFAP20 protein functions at the cilium inner junction (IJ), and is critical for connecting the component A- and B-microtubules together. The IJ is important in both motile and non-motile cilia function. When disrupted in zebrafish, cfap20 mutants displayed defective heart patterning and spine development (linked to motile cilia), and progressive retinal dystrophy and vision loss (linked to non-motile cilia). Mutations in the worm (C. elegans) Cfap-20 gene caused learning deficits resulting from dysfunction of non-motile cilia on sensory neurons. The researchers then showed that Cfap-20 loss caused an “open-seam” phenotype where the IJ was not connected, and the tubules were splayed apart. In the photoreceptor cells this led to shedding of parts of the cell and eventual cell death. These results show that CFAP20 bridges the two disparate cilia categories, and plays many important roles in cells of various organisms.
In conjunction, this group identified CFAP20 variants as a novel cause of blindness in humans. Retinitis pigmentosa (RP) is the most common form of inherited retinal dystrophy, with visual deficit often starting in childhood. Sufferers experience tunnel vision and early-onset blindness. Human sequencing identified 8 patients, from 4 families, with RP and CFAP20 variants predicted to be disease-causing. By introducing the human CFAP20 variants into their mutant zebrafish, Chrystal et al. showed that patient variants retained different levels of functionality, and this correlated with patient disease severity. The researchers also propose CFAP20 as a putative cause of other disorders involving cilia (infertility, mental disability, epilepsy which were all found in this patient cohort) arising from more damaging CFAP20 mutations. Functional testing demonstrated that some mutations were more damaging to function than others and could explain the variable levels of symptoms observed between patients. Additionally, functional testing has provided evidence that reintroducing the wildtype CFAP20 sequence can reverse symptoms, the strategy used in human gene therapy.
This work discovered a novel disease mechanism for retinitis pigmentosa originating at the inner junction, and could implicate other proteins that function at the IJ. While RP is the most common form of inherited retinal dystrophy, only one therapeutic currently exists, Luxturna, a gene-specific therapy to mutations in a gene called RPE65. This paper shows that CFAP20 is a novel cause of RP, adding to the molecular genetic underpinnings of the condition. Furthermore, these studies have provided 4 families with a genetic diagnosis that will allow for subsequent genetic counselling and intervention.
This project was a hugely collaborative effort, with thirty listed authors. A major strength of this project was the establishment of both national (University of Alberta, Simon Fraser University, University of Calgary) and international (University College London, Moorfields Hospital) collaboration. These collaborations allowed for the researchers to combine human genetics and clinical reporting, C. elegans and zebrafish disease modelling, and a mammalian cell culture model that strengthened the conclusions. While these groups had no previous history of collaboration, they have now developed strong working relationships and hope to collaborate again in the future.
About Paul William Chrystal
Paul William Chrystal completed his PhD in developmental genetics at Newcastle University, UK before moving to Edmonton, Canada for postdoctoral studies. He led this study in the laboratory of Dr. W. Ted Allison at the University of Alberta, completing the zebrafish disease modelling, and contributing to analysis, and manuscript preparation. Dr. Chrystal is passionate about understanding the genetics of inherited retinal diseases and translating this knowledge into patient therapeutics. He currently works at the University of Toronto, Canada as a postdoctoral fellow testing gene therapy approaches for preservation of sight in Usher’s syndrome.
Sources of funding
This project was funded by the Canadian Institutes of Health Research (CIHR; grants PJT-156042 and MOP-142243), CIHR Rare Disease Models & Mechanisms award (M-UBC-27R00211), and the Natural Sciences and Engineering Research Council of Canada (NSERC; grant RGPIN-2019-04825). Student grants provided by NSERC, Alberta Innovates and CIHR. The UK team were supported by Fight For Sight UK (5045/46), National Institute of Health Research Biomedical Research Centre (NIHRBRC), NIHR-BRC Moorfields Eye Charity (Stephen and Elizabeth Archer in memory of Marion Woods), Wellcome Trust (206619/Z/17/Z), Science Foundation Ireland in partnership with BBSRC (16/BBSRC/3394), and Imperial Health Charity. Additional support provided by Olive Young Fund, University Hospital Foundation, Women and Children’s Health Research Institute Innovation Grant (WCHRI 2846), and the Michael Smith Foundation for Health Research.
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